Bt maize expressing Cry1Ab was approved for release in South Africa for control of Busseola fusca (Fuller) (Lepidoptera: Noctuidae) and Chilo partellus (Swinhoe) (Lepidoptera: Crambidae) in 1998. During 2012, a stacked Bt maize event, expressing Cry2Ab2 + Cry1A.105, was also approved for control of these abovementioned pests. The aim of this study was to determine the effects of two Bt maize events expressing Cry1Ab (MON810 and Bt11) and a Bt maize event expressing Cry2Ab2 + Cry1A.105 (MON89034) on selected lepidopteran non-target pest species and certain lepidopteran stem borer species in South Africa. Results from previous studies and this study will provide information regarding efficacy of Bt maize against pests which have not been evaluated yet. Insects with significance in maize agro-ecosystems in South Africa as well as the rest of Africa, were prioritised and laboratory bioassays were conducted to evaluate the effect of Bt maize against these selected target and non-target pest species. Studies were conducted on three stem borers C. partellus, Eldana saccharina and Sesamia calamistis and three nontarget lepidopteran pest species Agrotis segetum, Helicoverpa armigera and Spodoptera exempta. Results showed that MON810 maize was not effective against A. segetum larvae feeding on maize seedlings. Differential levels of survival were observed between two A. segetum populations on MON89034 with a population from Polokwane showing survival on the stacked maize event. No S. exempta 1st instar larvae survived on MON810 and MON89034 but 3rd instar larvae survived on MON810 maize. Helicoverpa armigera larvae survived on ears of MON810 maize plants but not on events MON89034 or Bt 11. Results further indicated that C. partellus larvae were highly susceptible to these three Bt maize events. Larval survival of S. calamistis was recorded for larvae feeding on MON810 and Bt11 maize ears but not on MON89034 maize ears. Bt maize during the vegetative growth stages therefore effectively controlled C. partellus, S. calamistis and E. saccharina but the latter two species was not effectively controlled when feeding on ear tissue. This study provides important information on the effects of Bt maize on the most important non-target pest species of maize in sub-Saharan Africa. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

Bt maize expressing Cry1Ab was approved for release in South Africa for control of Busseola fusca (Fuller) (Lepidoptera: Noctuidae) and Chilo partellus (Swinhoe) (Lepidoptera: Crambidae) in 1998. During 2012, a stacked Bt maize event, expressing Cry2Ab2 + Cry1A.105, was also approved for control of these abovementioned pests. The aim of this study was to determine the effects of two Bt maize events expressing Cry1Ab (MON810 and Bt11) and a Bt maize event expressing Cry2Ab2 + Cry1A.105 (MON89034) on selected lepidopteran non-target pest species and certain lepidopteran stem borer species in South Africa. Results from previous studies and this study will provide information regarding efficacy of Bt maize against pests which have not been evaluated yet. Insects with significance in maize agro-ecosystems in South Africa as well as the rest of Africa, were prioritised and laboratory bioassays were conducted to evaluate the effect of Bt maize against these selected target and non-target pest species. Studies were conducted on three stem borers C. partellus, Eldana saccharina and Sesamia calamistis and three nontarget lepidopteran pest species Agrotis segetum, Helicoverpa armigera and Spodoptera exempta. Results showed that MON810 maize was not effective against A. segetum larvae feeding on maize seedlings. Differential levels of survival were observed between two A. segetum populations on MON89034 with a population from Polokwane showing survival on the stacked maize event. No S. exempta 1st instar larvae survived on MON810 and MON89034 but 3rd instar larvae survived on MON810 maize. Helicoverpa armigera larvae survived on ears of MON810 maize plants but not on events MON89034 or Bt 11. Results further indicated that C. partellus larvae were highly susceptible to these three Bt maize events. Larval survival of S. calamistis was recorded for larvae feeding on MON810 and Bt11 maize ears but not on MON89034 maize ears. Bt maize during the vegetative growth stages therefore effectively controlled C. partellus, S. calamistis and E. saccharina but the latter two species was not effectively controlled when feeding on ear tissue. This study provides important information on the effects of Bt maize on the most important non-target pest species of maize in sub-Saharan Africa. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

Presented study compares impacts of the GM maize MON 88017 and the non-transgenic maize on the non-target invertebrates. The Diabrotica-resistant and herbicide-tolerant GM maize may become important for most of Europe as the spred of Diabrotica virgifera virgifera LeConte continues but its implementation has been hindered by fears of undesirable environmental effects. I confirmed that MON88017 expresses the insecticidal Cry3Bb1 toxin but I found no evidence of any negative impact on the monitored species. Both the composition and the species abundances of invertebrate communities were similar to those in the stands of the conventional non-transgenic maize varieties.

Genetically modified (GM) crops were introduced in South Africa in 1989 and commercially available by 1998. Legislation to control the use of GM crops was only implemented in 1999, with the genetically modified organisms (GMO) act (15 of 1999). In 2012 2.9 million ha of GM crops were planted in South Africa alone. GM Crops, such as Bt maize, are promoted as safer for the environment since no chemical pesticides are needed. However, recently GM crops have been making headlines as more and more studies find adverse effects of these crops on non-target organisms. The effects on aquatic environments have not yet been fully determined, even though traces of Bt residue have been found in water systems surrounding agricultural lands. The aim of this study was to establish the effects of the Bt toxin on fecundity, development and growth of Bulinus tropicus, a freshwater snail. The experiment made use of a static renewal tests to expose B. tropicus to 50 cm2 Bt maize and cotton leaves in 900 ml of synthetic freshwater. The snails were exposed for the duration of one full life cycle (embryo to adult). Endpoints measured included the development, growth, fecundity, and deformities of the reproductive organs. The results obtained showed retarded development and low embryo survival when the snails were exposed to cotton leaves, irrespective of the presence or absence of Bt, indicating to the possibility of trace residues of chemical pesticides may have been present on the leaves. Initial stimulated growth of hatchlings was observed for both Bt cotton and maize exposures, but after sexual maturity has been reached, ‘surplus’ energy was probably shared between growth and fecundity, resulting in a reduction of growth rate. Energy is gained from their diet, thus a sub-optimal diet would result in less energy available to functions such as growth and fecundity. Signs of developmental instability were found in the formation of the shell opening of the snails exposed to Bt. Fecundity decreased significantly after snails had been exposed to Bt maize / cotton leaves. No differences were found in the penis sheath-preputium length ratio, indicating that Bt had no deleterious effects on the reproductive organs. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2014

The evolution of pest resistance to Cry proteins threatens the long-term use of Bt crops. Busseola fusca developed resistance to Bt maize in South Africa but the mechanism of resistance is not well understood. According to the gut microbiota theory, extensive cell lysis caused by Cry proteins provide gut microbes access to the more favourable environment of the hemocoel where they germinate and reproduce, causing septicemia and death of the host. This theory brought on questions about the role of gut microbes in the efficacy of Bt maize against target lepidopteran pests. The aim of this study was to determine whether microbes present in the mid-gut of B. fusca influence the efficacy of Cry 1Ab proteins. Larvae were collected from 30 different geographical locations, dissected to excise the midgut and mid-gut content which was separated according to morphological types. The morphological types were used to test the antibiotic susceptibility of the bacteria and proved that ciprofloxacin, ampicillin and doxycycline were the most effective bacteriostatic and bactericidal antibiotics. These three antibiotics were exposed to the morphological types at different concentrations to visualise the possible deleterious effects of the antibiotics on the bacteria. This visualisation was performed by observing the growth curve of the bacteria in the presence of the combination of antibiotics. The antibiotics concentration of 500 μg/ml showed the highest efficacy compared to the other concentrations tested. An antibiotic concentration of 500 μg/ml of ciprofloxacin, ampicillin and doxycycline was incorporated into an artificial diet for the larvae to feed on for 7 days. This method was used to rid the larvae of gut microbes before allowing them to feed on Bt maize (MON810) plant material expressing Cry proteins. The results suggests that by placing antibiotic reared larvae on a Bt plant, the absence of the mid-gut microbes contributed to larvae survival on Bt maize. This observation will contribute to understanding the role of gut microbes on the efficacy of Cry proteins. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

Genetically modified (GM) crops were introduced in South Africa in 1989 and commercially available by 1998. Legislation to control the use of GM crops was only implemented in 1999, with the genetically modified organisms (GMO) act (15 of 1999). In 2012 2.9 million ha of GM crops were planted in South Africa alone. GM Crops, such as Bt maize, are promoted as safer for the environment since no chemical pesticides are needed. However, recently GM crops have been making headlines as more and more studies find adverse effects of these crops on non-target organisms. The effects on aquatic environments have not yet been fully determined, even though traces of Bt residue have been found in water systems surrounding agricultural lands. The aim of this study was to establish the effects of the Bt toxin on fecundity, development and growth of Bulinus tropicus, a freshwater snail. The experiment made use of a static renewal tests to expose B. tropicus to 50 cm2 Bt maize and cotton leaves in 900 ml of synthetic freshwater. The snails were exposed for the duration of one full life cycle (embryo to adult). Endpoints measured included the development, growth, fecundity, and deformities of the reproductive organs. The results obtained showed retarded development and low embryo survival when the snails were exposed to cotton leaves, irrespective of the presence or absence of Bt, indicating to the possibility of trace residues of chemical pesticides may have been present on the leaves. Initial stimulated growth of hatchlings was observed for both Bt cotton and maize exposures, but after sexual maturity has been reached, ‘surplus’ energy was probably shared between growth and fecundity, resulting in a reduction of growth rate. Energy is gained from their diet, thus a sub-optimal diet would result in less energy available to functions such as growth and fecundity. Signs of developmental instability were found in the formation of the shell opening of the snails exposed to Bt. Fecundity decreased significantly after snails had been exposed to Bt maize / cotton leaves. No differences were found in the penis sheath-preputium length ratio, indicating that Bt had no deleterious effects on the reproductive organs. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2014

The evolution of pest resistance to Cry proteins threatens the long-term use of Bt crops. Busseola fusca developed resistance to Bt maize in South Africa but the mechanism of resistance is not well understood. According to the gut microbiota theory, extensive cell lysis caused by Cry proteins provide gut microbes access to the more favourable environment of the hemocoel where they germinate and reproduce, causing septicemia and death of the host. This theory brought on questions about the role of gut microbes in the efficacy of Bt maize against target lepidopteran pests. The aim of this study was to determine whether microbes present in the mid-gut of B. fusca influence the efficacy of Cry 1Ab proteins. Larvae were collected from 30 different geographical locations, dissected to excise the midgut and mid-gut content which was separated according to morphological types. The morphological types were used to test the antibiotic susceptibility of the bacteria and proved that ciprofloxacin, ampicillin and doxycycline were the most effective bacteriostatic and bactericidal antibiotics. These three antibiotics were exposed to the morphological types at different concentrations to visualise the possible deleterious effects of the antibiotics on the bacteria. This visualisation was performed by observing the growth curve of the bacteria in the presence of the combination of antibiotics. The antibiotics concentration of 500 μg/ml showed the highest efficacy compared to the other concentrations tested. An antibiotic concentration of 500 μg/ml of ciprofloxacin, ampicillin and doxycycline was incorporated into an artificial diet for the larvae to feed on for 7 days. This method was used to rid the larvae of gut microbes before allowing them to feed on Bt maize (MON810) plant material expressing Cry proteins. The results suggests that by placing antibiotic reared larvae on a Bt plant, the absence of the mid-gut microbes contributed to larvae survival on Bt maize. This observation will contribute to understanding the role of gut microbes on the efficacy of Cry proteins. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

Resumo: Comercialmente os híbridos de milho transgênico são hemizigotos para o evento que os define, ou seja, das linhagens parentais cruzadas, uma é transgênica e a outra convencional. No entanto, híbridos homozigotos também podem ser obtidos pelo do cruzamento de duas linhagens parentais transgênicas. Assim, uma vez que a influência de genes exógenos é pouco conhecida na morfologia, anatomia e fertilidade masculina de híbridos de milho, buscamos com esse trabalho verificar a influência do número de alelos transgênicos nestes caracteres. Foram utilizados cinco diferentes híbridos com o mesmo evento (TC1507xMON89034xNK603) e o isogênico convencional de um deles, além de um híbrido com três diferentes eventos (Bt11, MIR162 e a piramidação de ambos) e seu isogênico convencional. Totalizando 18 tratamentos, que foram conduzidos em casa de vegetação, elaborado em delineamento de blocos casualizados com duas repetições. Quando as plantas estavam em V3-V4 duas delas foram extraídas para avaliação dos caracteres anatômicos do colmo, as restantes foram conduzidas até V5-V6 e avaliados os caracteres anatômicos da folha e raiz, morfológicos da raiz, matéria fresca e matéria seca. Para a avaliação dos caracteres de fertilidade masculina foram instalados em campo o híbrido com os três diferentes eventos, com cinco repetições e utilizadas três plantas como parcela útil. No terceiro dia após a antese a viabilidade e a germinação desses grãos de pólen foram avaliados. Todos os híbridos foram difere... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Commercially the transgenic maize hybrids are hemizygous for the event that defines them, that is, for crossed parental lines, one is transgenic and the other conventional. However, homozygous hybrids can also be obtained by crossing two transgenic parent lines. Thus, since the influence of exogenous genes is not well known in the morphology, anatomy and male fertility of maize hybrids, we seek to evaluate the influence of the number of transgenic alleles in these traits. Five different hybrids were used with the same event (TC1507xMON89034xNK603) and the conventional isogenic of one of them, besides a hybrid with three different events (Bt11, MIR162 and pyramidation of both and its conventional isogenic. Totaling 18 treatments, they were conducted in greenhouse, elaborated in a randomized complete block design with two replicates. When the plants were in V3-V4, two of them were extracted to evaluate the anatomical traits of the stem, the remaining were conducted to V5-V6 then evaluated the anatomical traits of leaf and root and morphology of root, fresh matter and dry matter. For the evaluation of the male fertility traits, the hybrids with three events different, were installed in the field, with five replications and three plants were used as a useful plot. On the third day after the anthesis the viability and germination of these grains were evaluated. All hybrids were different in the anatomical traits, the morphological components and the viability of the pollen grain d... (Complete abstract click electronic access below) / Mestre

A utilização de plantas transgênicas expressando proteínas inseticidas de Bacillus thuringiensis (Bt) tem sido a principal estratégia para o controle da lagarta-do-cartucho Spodoptera frugiperda (J.E. Smith) no Brasil. A resistência de S. frugiperda à proteína Vip3Aa20 foi recentemente isolada e caracterizada em condições de laboratório, a partir de uma população proveniente de Correntina-BA. Para subsidiar programas de Manejo da Resistência de Insetos (MRI), foram realizados estudos para entender a distribuição geográfica da resistência de S. frugiperda à proteína Vip3Aa20 no Brasil, mediante uso de métodos fenotípicos (proteína purificada e folhas de milho Bt) e genotípicos (F1 e F2 screen), além da elucidação da presença custo adaptativo associado à resistência a partir de uma linhagem resistente quase-isogênica. Para o monitoramento com proteína purificada, as médias de sobrevivência na dose discriminatória de 3600 ng Vip3Aa20/cm2 para 10 populações/safra de S. frugiperda das principais regiões produtoras de milho do Brasil, tiveram um aumento em número, no decorrer das safras, para as populações que diferiram da sobrevivência da linhagem SUS, por não sobreposição do I.C. (95%). As larvas sobreviventes deste monitoramento morreram quando transferidas para folhas de milho Bt expressando Vip3Aa20. Não houve sobreviventes no monitoramento da resistência utilizando folhas de milho Bt, para todas as populações avaliadas. As médias de frequência alélica da resistência estimadas pelo método F1 screen foram de 0,0069, 0,0051 e 0,0041 para as populações avaliadas na 2ª safra/2016, entressafra/2016 e 1ª safra/2017 respectivamente. Pelo método F2 screen, as médias de frequência alélica foram de 0,0030, 0,0036, 0,0054 e 0,0042 para as populações da 2ª safra/2016, entressafra/2016, 1ª safra/2017 e 2ª safra/2017 respetivamente. Foram selecionadas 3 linhagens resistentes a Vip3Aa20 pelo método F2 screen, a partir de populações provenientes de Peabiru-PR, São João da Boa Vista-SP e Casa Branca-SP. Pelo teste de complementaridade, o mesmo alelo da resistência que havia sido inicialmente isolado da região de Correntina-BA, foi verificado nestas novas linhagens resistentes selecionadas. Não foi verificada a presença de custo adaptativo associado à resistência de S. frugiperda a Vip3Aa20, além de um maior valor adaptativo de indivíduos heterozigotos mediante avaliação de parâmetros de tabela de vida e fertilidade. Os resultados do presente trabalho comprovaram que o alelo da resistência de S. frugiperda para a proteína Vip3Aa20 está amplamente distribuído nas principais regiões produtoras de milho no Brasil. Este fato aliado à ausência de custo adaptativo da resistência reforçam a necessidade de implementação de estratégias eficientes de MRI para retardar a evolução da resistência de S. frugiperda à Vip3Aa20 no Brasil. / The use of transgenic plants expressing Bacillus thuringiensis (Bt) insecticidal proteins has been the main strategy to control the fall armyworm Spodoptera frugiperda (J.E. Smith) in Brazil. The resistance of S. frugiperda to Vip3Aa20 protein was recently isolated and characterized in laboratory conditions from a Correntina-BA population. In order to subsidize Insect Resistance Management (IRM) programs, studies were carried out to understand the geographical distribution of S. frugiperda resistance to Vip3Aa20 protein in Brazil, through the use of phenotypic (purified protein and Bt maize leaves) and genotypic methods (F1 and F2 screen), in addition, the elucidation of fitness-cost presence associated with resistance from a near-isogenic resistant strain. For the purified protein monitoring, the mean survival at the diagnostic concentration of 3,600 ng Vip3Aa20/cm2 for 10 populations/season of S. frugiperda from the main maize producing regions of Brazil, had an increase in the number of populations with significant difference by non-overlap in the confidence interval (95 %) in relation with our reference of susceptibility. The surviving larvae of this monitoring died when transferred to leaves of Bt maize expressing Vip3Aa20. There were no survivors in resistance monitoring using Bt maize leaves for all populations evaluated. The mean frequency of resistance alleles estimated by the F1 screen method were 0.0069, 0.0051 and 0.0041 for the populations evaluated in the 2nd/2,016, the offseason/2,016 and the 1st/2,017 crop seasons, respectively. By the F2 screen method, the means of allele frequency were 0.0030, 0.0036, 0.0054 and 0.0042 for the populations of the 2nd/2,016, offseason/2,016, 1st/2,017 and 2nd/2,017 crop seasons, respectively. Three Vip3Aa20 resistant strains were selected by the F2 screen method from populations from Peabiru-PR, São João da Boa Vista-SP and Casa Branca-SP. By the complementarity test, the same resistance allele that had been initially isolated from the Correntina-BA region was verified in these new selected resistant strains. The presence of the adaptive cost associated to the resistance of S. frugiperda to Vip3Aa20 was not verified, besides a higher adaptive value of heterozygous individuals through evaluation of life table parameters and fertility. The results of the present work proved that the resistance allele of S. frugiperda for the Vip3Aa20 protein is widely distributed in the main maize producing regions in Brazil. This fact, coupled with the lack of adaptive cost of resistance, reinforces the need to implement efficient IRM strategies to delay the evolution of S. frugiperda resistance to Vip3Aa20 protein in Brazil.

<p>The western corn rootworm (WCR) <i>Diabrotica virgifera virgifera</i> Leconte is a major pest of maize in the United States. Currently, it is principally managed using Bt maize hybrids and neonicotinoid seed treatments (NSTs), which were concurrently introduced in the early 2000s. This simultaneous release, and subsequent rapid adoption, created a situation in which Bt maize hybrids were never assessed in the absence of NSTs, and vice versa. Consequently, neonicotinoids’ influence on refuge function, primarily whether these insecticides aid or hinder the production of a sufficient population of susceptible beetles to delay resistance, has not been assessed. Moreover, a mounting suite of detrimental environmental effects of NSTs have been documented, lending some urgency to questions about their necessity.</p><p>To determine the influence of NSTs on refuge function, untreated and treated Bt maize fields were planted with 5% untreated refuge marked with ¹⁵N. Throughout the field seasons of 2019 and 2020, adults were collected from these fields and analyzed to determine their natal host plant. Results documented that the numbers of refuge beetles produced by the 5% seed blend are likely insufficient to result in rates of mating to delay resistance development.</p><p>To determine if the effect of using NSTs in combination with Bt maize hybrids is additive, synergistic, or neutral at managing secondary soil pests in the Midwest, four 16 block fields were planted in 2018, 2019 and 2020 comparing four treatments (1. Untreated, Bt seed; 2. NST, Bt seed; 3. Untreated, non-Bt seed; 4. NST, non-Bt seed). Compact method sampling, root rating, and yield were used to document the presence of secondary soil pests. All three sampling years documented low abundance of white grubs and wireworms. There was minimal influence of NSTs on maize yield (2018: p = 0.07; 2019: p = 0.62; 2020: p = 0.056) and root damage (0-3 scale) (2018: M = 0.0092; 2019: M = 0.0091; 2020: M = 0.0361). These same fields were used to document the presence of NSTs in soil, as well as residues in nearby waterways. Results documented greatest soil and water clothianidin levels earlier in the season and declined as the season progressed.</p><p><br></p>